Platelet-activating factor (PAF) is a phospholipid which induces platelet aggregation and triggers the release reaction by a mechanism that is different from those of both adenosine diphosphate (ADP) and arachidonic acid (AA). Recent published data suggest that endogenous PAF from platelets may account for that part of thrombin, collagen and ionophore A23187 induced aggregation which is not prevented by blocking the ADP and AA mechanisms, but which is diminished by phospholipase A2 inhibitors. Thus a clear understanding of its mode of formation, its effects on platelets and other cells and its subsequent metabolism should lead to the development of rationales for the prevention of thrombotic episodes. The specific aims of this proposal are (1) to determine whether an arachidonic acid containing alkyl ether phospholipid is present in human platelets and if so, how it is changed in response to thrombin and collagen, (2) to determine basal levels of lyso-PAF in human platelets by its chemical conversion of PAF and quantitation by radioimmunoassay after purification by HPLC, (3) to develop a radioimmunoassay for PAF, (4) to evaluate the platelet aggregating activity of PAF analogs containing saturated and unsaturated fatty acids at 2-position of PAF.